Bacteriolysins, including phage-encoded lysins and bacteriocins are enzymes that target one of the four major cell wall peptidoglycan bonds leading to lysis of the bacterial cell wall. Lysins accumulate in the cytoplasm of infected bacteria during phage life cycle. At a genetically hard-coded time, another phage protein named holin is inserted in the cytoplasmic membrane leading to membrane disruption [1] enabling the lysin to access the peptidoglycan, thereby causing cell lysis and release of progeny phage [2]. Interestingly, exogenously applied recombinant lysins are able to cleave the integral peptidoglycan bonds of susceptible bacteria [3]. Non-phage bacteriolysins have been also described that are encoded by specific bacterial species and serve the purpose of lysing competing strains or autolysis at the septal region of the bacterial cell during cell division. Lysostaphin is a bacteriocin secreted by Staphylococcus simulans biovar staphylolyticus and directed against the cell wall of competing S. aureus [4]. The S. aureus autolysin (Atl) is a 138-kDa protein which is processed into two major peptidoglycan hydrolases and localizes on the cell wall at the septal region of an upcoming cell division site and plays a key role in cell separation [5]. Species-specific phages [6] and lysostaphin have been tested as potential therapeutics in numerous in vitro and in vivo studies [4, 7-10].
Phage lysins, lysostaphin, and autolysins all consist of a cell wall targeting (CWT) domain (or also known as cell wall binding domain; CBD) and one or more catalytic domains [4, 10-13]. CWTs recognize specific surface moieties, positioning the catalytic domain on the bacterial surface for enzymatic cleavage of the peptidoglycan. The catalytic domains are relatively conserved whereas CWTs are not conserved across species and impart species-specificity and can have variable binding affinities to different strains. Several studies reported the construction of truncated [8] or chimeric versions of phage lysins [7] showing that individual domains function when grafted into heterologous contexts such as green fluorescence protein (GFP). Lysostaphin CWT (GFP-CWT) was shown to bind to S. aureus as well as P. sacculi [9]. Similarly, GFP fusion of autolysin CWT binds to S. aureus with a dissociation constant of 15 nM [14]. Crystal structure studies show that Isolated CWTs have a compact structure and can fold independently. Thus, isolated CWT domain can be used to target heterologous proteins to the surface of bacteria at the site of infection.
Antibodies constitute the central pillar of the humoral immune response. Antibodies targeted against bacterial toxins capture and neutralize the toxins and as such can reduce the virulence of the respective bacterial species. However, antitoxin antibodies are not specifically targeted to the site of bacterial infection as they lack the ability to recognize bacterial cell wall. Antibodies that recognize bacterial surface antigens such as capsular polysaccharides and other surface moieties through their antigen binding Fab domains opsonize the bacteria allowing the innate immune response to bind and phagocytose the bacteria. This function is dependent on the Fc portion of the antibodies that undergo specific interaction with their receptors on the surface of phagocytic cells such as neutrophils or macrophages. The adaptive immune system requires prior encounter with the invading bacteria or a vaccine mimicking the respective bacteria to generate protective antibodies, a process that takes days to weeks before a fully functional protective response can be mounted. Therefore, active vaccination is useful after the onset of acute infections.